AUTOIMMUNE DIABETES: EARLY EVENTS IN ISLETS OF LANGERHANS

自身免疫性糖尿病：朗格汉斯岛的早期事件

• 批准号: 9197630
• 负责人: EMIL Raphael UNANUE
• 金额: $ 45.57万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-01 至 2020-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9197630
• 关键词: Acute; Address; Alleles; Antibodies; Area; Autoimmune Diabetes; Beta Cell; Binding; Blood; CD4 Positive T Lymphocytes; CRISPR/Cas technology; Cells; Charge; Chemotactic Factors; Chimera organism; Clinical; Complex; Defect; Dendritic Cells; Development; Diabetes Mellitus; Disease; Epitopes; Event; Experimental Models; Fatty acid glycerol esters; Flushing; Gatekeeping; Gene Expression Profile; Genetic; Genome; Grant; Human; ITGAX gene; Image; In Vitro; Inbred NOD Mice; Insulin; Insulin-Dependent Diabetes Mellitus; Investigation; Islets of Langerhans; Lymphocyte; Lymphoid; Migration Assay; Minor; Modeling; Monoclonal Antibodies; Movement; Mus; NOR Mouse; Pancreas; Peptide/MHC Complex; Peptides; Phase; Process; Publishing; Reagent; Recruitment Activity; Reporter; Reverse Transcriptase Polymerase Chain Reaction; Role; Site; T-Lymphocyte; Testing; Thymus Gland; Time; Tissues; Translating; XCL1 gene; XCR1 gene; autoreactivity; base; chemokine; chemokine receptor; congenic; imaging approach; in vivo; islet; lymph nodes; migration; public health relevance; receptor; transcription factor

 DESCRIPTION (provided by applicant): This proposal is based on a recently published study showing that a minor subset of dendritic cells, the CD11c+CD103+ DC, those under the control of the Batf3 transcription factor, are absolutely required for diabetes development in NOD mice. These DCs are found both in islets and pancreatic lymph nodes and act as gate-keepers for diabetes initiation. Our aims center on discovering the processes whereby CD103+DC localize in islets. We combine examination of chemokines, antibodies, and live imaging with new preliminary findings that point to NOR mice having a defect in islets CD103+ DC, but not in pLNs. These areas of emphasis may point to strategies that may be translated to human T1D. Aim 1 investigates the mechanism of CD103+ DC entry into islets. The role of chemokines in CD103+ DC recruitment will be examined both in vitro and in vivo. Chemokine blockade will be tested in vivo. In vitro migration assays will define chemokines with the potential to recruit NOD-derived CD103+ DC to islets. RT- PCR will be used to determine which chemokines exist in islets of NOD mice at different times. Aim 2 will generate and examine several reagents intended to deplete or trace CD103+ DC in the NOD mouse. We have just generated an Xcr1-/- mice using Crispr/Cas technology which should bring definitive information on the role of this chemokine receptor in the islet localization of the CD103+ DCs. We will follow the same approaches used to examine the Batf3-/- mice just published. In a second subaim, a NOD.Xcr1-mOrange reporter mouse will be generated and used to trace the movement of CD103+ DC in the pancreas using live imaging approaches. We want to examine the traffic of CD103+DC in the pancreas and determine whether it may be the cell that transmigrates to the pLN. Finally an anti-Xcl1 monoclonal antibodies will be tested for its effects on CD103+ DC migration and protection from diabetes. Aim 3 takes advantage of our recent finding that NOR mice, which share 88% of their genome with NOD, lack the intra islet CD103+ DC but have them in LNs. By comparing the Batf3-/- mice with the NORs we expect to understand the mechanisms controlling early entrance of cells into islets. We examine this strain, and substrains containing NOR-derived alleles, to address the genetic behind the CD103+ DC entry into islets.

 描述（由应用程序提供）：该提案基于最近发表的一项研究，该研究表明，在BATF3转录因子控制下的树突状细胞CD11C+ CD103+ DC的一小部分是NOD小鼠中糖尿病发育绝对必需的。这些DC在胰岛和胰腺淋巴结中都发现，并充当糖尿病启动的守门人。我们的目标是发现CD103+DC本地化在胰岛中的过程。我们将趋化因子，抗体和实时成像的检查与新的初步发现相结合，该发现指出了在胰岛CD103+ DC中具有缺陷的NOR小鼠，但在PLN中却没有。这些强调的领域可能会指出可以转化为人类T1D的策略。 AIM 1研究了CD103+ DC进入胰岛的机制。趋化因子在CD103+ DC募集中的作用将在体外和体内检查。趋化因子封锁将在体内进行测试。体外迁移分析将定义趋化因子，并可能将点头衍生的CD103+ DC募集到胰岛。 RT-PCR将用于确定在不同时间的NOD小鼠胰岛中存在哪些趋化因子。 AIM 2将生成并检查旨在在NOD小鼠中复制或跟踪CD103+ DC的几种试剂。我们刚刚使用CRISPR/CAS技术生成了XCR1 - / - 小鼠，该小鼠应该为该趋化因子受体在CD103+ DC的胰岛定位中的作用带来明确的信息。我们将遵循用于检查刚刚发布的BATF3 - / - 小鼠的相同方法。在第二个subiaim中，将生成NOD.XCR1-MORANGE记者鼠标，并使用实时成像方法在胰腺中追踪CD103+ DC的运动。我们想检查胰腺中CD103+DC的流量，并确定是否可能是传输到PLN的细胞。最后，将测试抗XCL1单克隆抗体对CD103+ DC迁移的影响，并保护其免受糖尿病的保护。 AIM 3利用了我们最近的发现，即与点头共享基因组的88％的NOR小鼠缺乏Islet CD103+ DC，但将它们放在LNS中。通过将BATF3 - / - 小鼠与NORS进行比较，我们希望了解控制细胞早期入口胰岛的机制。我们检查了这种菌株和含有非源性等位基因的底座，以解决CD103+ DC进入胰岛的遗传。